Dr. Xuelian Luo received her Ph.D. and postdoctoral training in protein structure determination by nuclear magnetic resonance (NMR) spectroscopy. She is currently an Instructor in Dr. Jose Rizo-Rey's lab at UT Southwestern Medical Center. Her long-term goal as an independent scientist is to combine structural biology and cell biology to study cancer-related topics such as cell division and signal transduction. During her mentored training period, she will continue to study the structure and function of the Mad2 spindle checkpoint protein. UT Southwestern is a top biomedical institution with excellent reputations in both structural and cancer biology, and thus provides an excellent environment for her training. The Mad2 spindle checkpoint protein ensures the accurate separation of sister-chromatids by inhibiting the ubiquitin ligase activity of the anaphase-promoting complex (APC) until all chromatids achieve bipolar attachment to the mitotic spindle. Mad2 binds to and sequesters Cdc20, an activator of APC, thereby inhibiting APC. Dr. Luo showed that binding of Mad2 to Cdc20 requires a major conformational switch of Mad2. Remarkably, Mad2 undergoes a similarly dramatic structural change upon binding to its upstream regulator, Mad1. She now shows that, in the absence of ligands, apo-Mad2 spontaneously refolds into an "activated" structural state, which resembles the ligand-bound structure of Mad2 with a vacant ligand-binding site. This form of Mad2 is more potent in blocking APC activity. However, the unassisted structural transition of Mad2 occurs with a timescale of hours, suggesting that other checkpoint components, such as Mad1, might facilitate the conformational activation of Mad2 in vivo. Using a combination of biophysical, biochemical, and cell biological techniques, she proposes to investigate the mechanism and the functional consequences of the unusual structural malleability of Mad2. Aim 1 is to characterize the two folded states of Mad2 in the absence of ligands in vitro and in vivo. The focus of Aim 2 is to understand the mechanism by which Mad2 accomplishes the unusually large conformational change in vitro and in vivo. In Aim 3, she will investigate the kinetic folding pathways of Mad2 in vitro and the possibility of chaperone-assisted Mad2 folding in vivo. Malfunction of the spindle checkpoint may contribute to genetic instability and aneuploidy of tumor cells, and several anti-cancer drugs, including Taxol, kill cancer cells by activating this checkpoint. Her proposed studies will facilitate the design of new anti-cancer drugs.